Establishment Of Plant Regeneration System And Preliminary Studies On Genetic Transformation Of Rose

Rose (Rosa hybrida L.) is one kind of the most important ornamental plants, which is popular because of its beautiful color, shape and full-bodied fragrant, concurrent flowers and its flowers language. It is applied to landscaping in modern cities with a wide range because of its good ecological suitability. Because of narrow genetic base and traditional breeding methods need to take a long term of modern roses, which delays the development of rose breeding at present, bioengineering offers a new approach for breeding work, which can keep the other properties relative stability, just change the properties which are dominated by exogenous gene, and can use the genes of other organisms, which provides a new way for improving rose quality.An in vitro micropropagation system for three rose was established by discussing the factors that affected micropropagation. A plant regeneration system by indirect organogenesis was established for Rosa odorata var gigantea and Rosa chinensis ’Pallida’. Besides, the optimal conditions for Agrobacterium-mediated transformation of basal callus of three roses was preliminary discussed. The main results were as the following:1. An in vitro micropropagation system was established for three rose. The optimal medium for germination of R.odorata var gigantea and R.chinensis ’Pallida’ was: MS+6-BA1.5mg/L+NAA0.12mg/L. The optimal medium for germination of ’Mitang’ was:MS+6-BA1.0mg/L+NAA0.1mg/L. The optimal medium for shoot multiplication of R. odorata var gigantea was MS+6-BA1.0mg/L+NAA0.01mg/L+GA30.3mg/L. The optimal medium for shoot multiplication of R. chinensis ’Pallida’ was:MS+6-BA1.0mg/L+NAA0.01mg/L+GA30.2mg/L. The optimal medium for shoot multiplication of ’Mitang’ was:MS+6-BA1.0mg/L+NAA0.01mg/L+GA30.3mg/L. The optimal rooting mediums for three roses were1/2MS+NAA0.15mg/L. The survival rate in the open can be up to80%by domestication and transplanting.2. The optimal mediums for callus induction of three roses were got using leaflets as explant. The optimal medium for callus induction of R.odorata var gigantea was: MS+2,4-D4.0mg/L with the rate of induction93.33%. The optimal medium for callus induction of R.chinensis ’Pallida’ was:MS+2,4-D3.0mg/L, with the rate of induction94.16%. The optimal medium for callus induction of ’Mitang’ was:MS+2,4-D4.0mg/L and MS+2,4-D5.0mg/L,the rates of induction both reach100%with good state of growth.3. The differentiation of callus was affected by the combined action of medium component, cultural condition and medium additives. The plant regeneration system via indirect organogenesis was established for R. odorata var gigantea and R. chinensis ’Pallida’. Being cultured on differentiation medium (MS+TDZ3.0mg/L+GA31.0mg/L+NAA0.1mg/L) for15d in darkness, callus of R. odorata var gigantea showed the highest frequency of adventitious bud induction with18%. Regeneration plant can grow on multiplication medium. Being cultured on differentiation medium (MS+TDZ8.0mg/L+NAA0.1mg/L) for15d in darkness and transferred to MS+TDZ3.0mg/L+6-BA1.0mg/L+GA31.0mg/L+NAA0.1mg/L after two months, somatic embryogenesis can be induced from callus with the rate of13.33%. Being cultured on differentiation medium (MS+TDZ2.0mg/L+6-BA1.0mg/L+GA31.0mg/L+NAA0.1mg/L) for15d in darkness, R. chinensis ’Pallida’ showed the highest frequency of adventitious bud induction with18.67%. Being cultured on differentiation medium (MS+TDZ3.0mg/L+6-BA1.0mg/L+GA31.0mg/L+NAA0.1mg/L)for15d in darkness, somatic embryogensis can be induced from callus with the rate of10.67%. Being cultured on differentiation medium (MS+TDZ4.0mg/L) for15d in darkness,4. An Agrobacterium tumefaciens-mediated transformation protocol was established preliminary for three roses using callus as inoculation material. The optimal conditions for Agrobacterium tumefaciens-mediated transformation were investigated. The result showed that infected25min and co-cultured4d, the rate of GUS transient expression for R. odorata var gigantea can be up to87.5%. Being infected15min and co-culture5d, the rate of GUS transient expression for R. chinensis ’Pallida’ can be up to55.94%. Being infected15min and co-culture4d, the rate of GUS transient expression for ’Mitang’ can be up to63.34%.